Biogenesis of the plant secondary cell wall involves many important aspects, such as phenolic compound deposition and often silica encrustation. Previously, we demonstrated the importance of the exocyst subunit EXO70H4 for biogenesis of the trichome secondary cell wall, namely for deposition of the autofluorescent and callose-rich cell wall layer. Here, we reveal that EXO70H4-driven cell wall biogenesis is constitutively active in the mature trichome, but also can be activated elsewhere upon pathogen attack, giving this study a broader significance with an overlap into phytopathology. To address the specificity of EXO70H4 among the EXO70 family, we complemented the exo70H4-1 mutant by 18 different Arabidopsis (Arabidopsis thaliana) EXO70 paralogs subcloned under the EXO70H4 promoter. Only EXO70H4 had the capacity to rescue the exo70H4-1 trichome phenotype. Callose deposition phenotype of exo70H4-1 mutant is caused by impaired secretion of PMR4, a callose synthase responsible for the synthesis of callose in the trichome. PMR4 colocalizes with EXO70H4 on plasma membrane microdomains that do not develop in the exo70H4-1 mutant. Using energy-dispersive x-ray microanalysis, we show that both EXO70H4- and PMR4-dependent callose deposition in the trichome are essential for cell wall silicification.

Department of Experimental Plant Biology Faculty of Sciences Charles University Prague Czech Republic

Institute of Experimental Botany Academy of Sciences of the Czech Republic Prague Czech Republic

Institute of Scientific Instruments of the Academy of Sciences of the Czech Republic Brno Czech Republic

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Bischoff V, Nita S, Neumetzler L, Schindelasch D, Urbain A, Eshed R, Persson S, Delmer D, Scheible W-R (2010) TRICHOME BIREFRINGENCE and its homolog AT5G01360 encode plant-specific DUF231 proteins required for cellulose biosynthesis in Arabidopsis. Plant Physiol 153: 590–602 PubMed PMC

Blümke A, Somerville SC, Voigt CA (2013) Transient expression of the Arabidopsis thaliana callose synthase PMR4 increases penetration resistance to powdery mildew in barley. Adv Biosci Biotechnol 04: 810–813

Bonke M, Thitamadee S, Mähönen AP, Hauser M-T, Helariutta Y (2003) APL regulates vascular tissue identity in Arabidopsis. Nature 426: 181–186 PubMed

Brennwald P, Rossi G (2007) Spatial regulation of exocytosis and cell polarity: yeast as a model for animal cells. FEBS Lett 581: 2119–2124 PubMed PMC

Brugiére T, Exley C (2017) Callose-associated silica deposition in Arabidopsis. J Trace Elem Med Biol 39: 86–90 PubMed

Cai G, Faleri C, Del Casino C, Emons AMC, Cresti M (2011) Distribution of callose synthase, cellulose synthase, and sucrose synthase in tobacco pollen tube is controlled in dissimilar ways by actin filaments and microtubules. Plant Physiol 155: 1169–1190 PubMed PMC

Chen C, Liu M, Jiang L, Liu X, Zhao J, Yan S, Yang S, Ren H, Liu R, Zhang X (2014) Transcriptome profiling reveals roles of meristem regulators and polarity genes during fruit trichome development in cucumber (Cucumis sativus L.). J Exp Bot 65: 4943–4958 PubMed PMC

Chérif M, Menzies JG, Benhamou N, Bélanger RR (1992) Studies of silicon distribution in wounded and Pythium ultimum infected cucumber plants. Physiol Mol Plant Pathol 41: 371–385

Clough SJ, Bent AF (1998) Floral dip: a simplified method for Agrobacterium-mediated transformation of Arabidopsis thaliana. Plant J 16: 735–743 PubMed

Cvrčková F, Grunt M, Bezvoda R, Hála M, Kulich I, Rawat A, Zárský V (2012) Evolution of the land plant exocyst complexes. Front Plant Sci 3: 159. PubMed PMC

Dong X, Hong Z, Chatterjee J, Kim S, Verma DPS (2008) Expression of callose synthase genes and its connection with Npr1 signaling pathway during pathogen infection. Planta 229: 87–98 PubMed

Drakakaki G, van de Ven W, Pan S, Miao Y, Wang J, Keinath NF, Weatherly B, Jiang L, Schumacher K, Hicks G, et al. (2012) Isolation and proteomic analysis of the SYP61 compartment reveal its role in exocytic trafficking in Arabidopsis. Cell Res 22: 413–424 PubMed PMC

Drdová EJ, Synek L, Pečenková T, Hála M, Kulich I, Fowler JE, Murphy AS, Zárský V (2013) The exocyst complex contributes to PIN auxin efflux carrier recycling and polar auxin transport in Arabidopsis. Plant J 73: 709–719 PubMed

Elias M, Drdova E, Ziak D, Bavlnka B, Hala M, Cvrckova F, Soukupova H, Zarsky V (2003) The exocyst complex in plants. Cell Biol Int 27: 199–201 PubMed

Ellinger D, Glöckner A, Koch J, Naumann M, Stürtz V, Schütt K, Manisseri C, Somerville SC, Voigt CA (2014) Interaction of the Arabidopsis GTPase RabA4c with its effector PMR4 results in complete penetration resistance to powdery mildew. Plant Cell 26: 3185–3200 PubMed PMC

Ellinger D, Naumann M, Falter C, Zwikowics C, Jamrow T, Manisseri C, Somerville SC, Voigt CA (2013) Elevated early callose deposition results in complete penetration resistance to powdery mildew in Arabidopsis. Plant Physiol 161: 1433–1444 PubMed PMC

Exley C. (2015) A possible mechanism of biological silicification in plants. Front Plant Sci 6: 853. PubMed PMC

Fauteux F, Chain F, Belzile F, Menzies JG, Bélanger RR (2006) The protective role of silicon in the Arabidopsis-powdery mildew pathosystem. Proc Natl Acad Sci USA 103: 17554–17559 PubMed PMC

Fendrych M, Synek L, Pecenková T, Drdová EJ, Sekeres J, de Rycke R, Nowack MK, Zársky V (2013) Visualization of the exocyst complex dynamics at the plasma membrane of Arabidopsis thaliana. Mol Biol Cell 24: 510–520 PubMed PMC

Frye CA, Innes RW (1998) An Arabidopsis mutant with enhanced resistance to powdery mildew. Plant Cell 10: 947–956 PubMed PMC

Geldner N, Dénervaud-Tendon V, Hyman DL, Mayer U, Stierhof Y-D, Chory J (2009) Rapid, combinatorial analysis of membrane compartments in intact plants with a multicolor marker set. Plant J 59: 169–178 PubMed PMC

Ghanmi D, McNally DJ, Benhamou N, Menzies JG, Bélanger RR (2004) Powdery mildew of Arabidopsis thaliana: a pathosystem for exploring the role of silicon in plant–microbe interactions. Physiol Mol Plant Pathol 64: 189–199

Grefen C, Donald N, Hashimoto K, Kudla J, Schumacher K, Blatt MR (2010) A ubiquitin-10 promoter-based vector set for fluorescent protein tagging facilitates temporal stability and native protein distribution in transient and stable expression studies. Plant J 64: 355–365 PubMed

Guerriero G, Hausman J-F, Legay S (2016) Silicon and the plant extracellular matrix. Front Plant Sci 7: 463. PubMed PMC

Guseman JM, Lee JS, Bogenschutz NL, Peterson KM, Virata RE, Xie B, Kanaoka MM, Hong Z, Torii KU (2010) Dysregulation of cell-to-cell connectivity and stomatal patterning by loss-of-function mutation in Arabidopsis chorus (glucan synthase-like 8). Development 137: 1731–1741 PubMed

Heider MR, Gu M, Duffy CM, Mirza AM, Marcotte LL, Walls AC, Farrall N, Hakhverdyan Z, Field MC, Rout MP, et al. (2016) Subunit connectivity, assembly determinants and architecture of the yeast exocyst complex. Nat Struct Mol Biol 23: 59–66 PubMed PMC

Heider MR, Munson M (2012) Exorcising the exocyst complex. Traffic 13: 898–907 PubMed PMC

Hodson MJ. (2016) The development of phytoliths in plants and its influence on their chemistry and isotopic composition. Implications for palaeoecology and archaeology. J Archaeol Sci 68: 62–69

Hong D, Jeon BW, Kim SY, Hwang J-U, Lee Y (2016) The ROP2-RIC7 pathway negatively regulates light-induced stomatal opening by inhibiting exocyst subunit Exo70B1 in Arabidopsis. New Phytol 209: 624–635 PubMed

Hong Z, Delauney AJ, Verma DP (2001) A cell plate-specific callose synthase and its interaction with phragmoplastin. Plant Cell 13: 755–768 PubMed PMC

Huang L, Chen X-Y, Rim Y, Han X, Cho WK, Kim S-W, Kim J-Y (2009) Arabidopsis glucan synthase-like 10 functions in male gametogenesis. J Plant Physiol 166: 344–352 PubMed

Hülskamp M, Misŕa S, Jürgens G (1994) Genetic dissection of trichome cell development in Arabidopsis. Cell 76: 555–566 PubMed

Iglesias VA, Meins F Jr (2000) Movement of plant viruses is delayed in a beta-1,3-glucanase-deficient mutant showing a reduced plasmodesmatal size exclusion limit and enhanced callose deposition. Plant J 21: 157–166 PubMed

Jacobs AK, Lipka V, Burton RA, Panstruga R, Strizhov N, Schulze-Lefert P, Fincher GB (2003) An Arabidopsis callose synthase, GSL5, is required for wound and papillary callose formation. Plant Cell 15: 2503–2513 PubMed PMC

Jakoby MJ, Falkenhan D, Mader MT, Brininstool G, Wischnitzki E, Platz N, Hudson A, Hülskamp M, Larkin J, Schnittger A (2008) Transcriptional profiling of mature Arabidopsis trichomes reveals that NOECK encodes the MIXTA-like transcriptional regulator MYB106. Plant Physiol 148: 1583–1602 PubMed PMC

Kalmbach L, Hématy K, De Bellis D, Barberon M, Fujita S, Ursache R, Daraspe J, Geldner N (2017) Transient cell-specific EXO70A1 activity in the CASP domain and Casparian strip localization. Nat Plants 3: 17058. PubMed

Karimi M, Bleys A, Vanderhaeghen R, Hilson P (2007) Building blocks for plant gene assembly. Plant Physiol 145: 1183–1191 PubMed PMC

Kulich I, Pečenková T, Sekereš J, Smetana O, Fendrych M, Foissner I, Höftberger M, Zárský V (2013) Arabidopsis exocyst subcomplex containing subunit EXO70B1 is involved in autophagy-related transport to the vacuole. Traffic 14: 1155–1165 PubMed

Kulich I, Vojtíková Z, Glanc M, Ortmannová J, Rasmann S, Žárský V (2015) Cell wall maturation of Arabidopsis trichomes is dependent on exocyst subunit EXO70H4 and involves callose deposition. Plant Physiol 168: 120–131 PubMed PMC

Law C, Exley C (2011) New insight into silica deposition in horsetail (Equisetum arvense). BMC Plant Biol 11: 112. PubMed PMC

Leroux O, Leroux F, Mastroberti AA, Santos-Silva F, Van Loo D, Bagniewska-Zadworna A, Van Hoorebeke L, Bals S, Popper ZA, de Araujo Mariath JE (2013) Heterogeneity of silica and glycan-epitope distribution in epidermal idioblast cell walls in Adiantum raddianum laminae. Planta 237: 1453–1464 PubMed

Levy A, Erlanger M, Rosenthal M, Epel BL (2007) A plasmodesmata-associated β-1,3-glucanase in Arabidopsis. Plant J 49: 669–682 PubMed

Li S, van Os GMA, Ren S, Yu D, Ketelaar T, Emons AMC, Liu C-M (2010) Expression and functional analyses of EXO70 genes in Arabidopsis implicate their roles in regulating cell type-specific exocytosis. Plant Physiol 154: 1819–1830 PubMed PMC

Luo G, Zhang J, Guo W (2014) The role of Sec3p in secretory vesicle targeting and exocyst complex assembly. Mol Biol Cell 25: 3813–3822 PubMed PMC

Munson M, Novick P (2006) The exocyst defrocked, a framework of rods revealed. Nat Struct Mol Biol 13: 577–581 PubMed

Mylle E, Codreanu M-C, Boruc J, Russinova E (2013) Emission spectra profiling of fluorescent proteins in living plant cells. Plant Methods 9: 10. PubMed PMC

Neděla V, Tihlaříková E, Hřib J (2015) The low-temperature method for study of coniferous tissues in the environmental scanning electron microscope. Microsc Res Tech 78: 13–21 PubMed

Nielsen ME, Feechan A, Böhlenius H, Ueda T, Thordal-Christensen H (2012) Arabidopsis ARF-GTP exchange factor, GNOM, mediates transport required for innate immunity and focal accumulation of syntaxin PEN1. Proc Natl Acad Sci USA 109: 11443–11448 PubMed PMC

Oda Y, Iida Y, Nagashima Y, Sugiyama Y, Fukuda H (2015) Novel coiled-coil proteins regulate exocyst association with cortical microtubules in xylem cells via the conserved oligomeric golgi-complex 2 protein. Plant Cell Physiol 56: 277–286 PubMed

Pecenková T, Hála M, Kulich I, Kocourková D, Drdová E, Fendrych M, Toupalová H, Zársky V (2011) The role for the exocyst complex subunits Exo70B2 and Exo70H1 in the plant-pathogen interaction. J Exp Bot 62: 2107–2116 PubMed PMC

Peragine A, Yoshikawa M, Wu G, Albrecht HL, Poethig RS (2004) SGS3 and SGS2/SDE1/RDR6 are required for juvenile development and the production of trans-acting siRNAs in Arabidopsis. Genes Dev 18: 2368–2379 PubMed PMC

Perry CC, Williams RJP, Fry SC (1987) Cell wall biosynthesis during silicification of grass hairs. J Plant Physiol 126: 437–448

Picco A, Irastorza-Azcarate I, Specht T, Böke D, Pazos I, Rivier-Cordey A-S, Devos DP, Kaksonen M, Gallego O (2017) The in vivo architecture of the exocyst provides structural basis for exocytosis. Cell 168: 400–412.e18 PubMed

Robinson NGG, Guo L, Imai J, Toh-E A, Matsui Y, Tamanoi F (1999) Rho3 of Saccharomyces cerevisiae, which regulates the actin cytoskeleton and exocytosis, is a GTPase which interacts with Myo2 and Exo70. Mol Cell Biol 19: 3580–3587 PubMed PMC

Russo VM, Bushnell WR (1989) Responses of barley cells to puncture by microneedles and to attempted penetration by Erysiphe graminis f.sp. hordei. Can J Bot 67: 2912–2921

Sabol P, Kulich I, Žárský V (2017) RIN4 recruits the exocyst subunit EXO70B1 to the plasma membrane. J Exp Bot 68: 3253–3265 PubMed PMC

Saedler R, Mathur N, Srinivas BP, Kernebeck B, Hülskamp M, Mathur J (2004) Actin control over microtubules suggested by DISTORTED2 encoding the Arabidopsis ARPC2 subunit homolog. Plant Cell Physiol 45: 813–822 PubMed

Samuel MA, Chong YT, Haasen KE, Aldea-Brydges MG, Stone SL, Goring DR (2009) Cellular pathways regulating responses to compatible and self-incompatible pollen in Brassica and Arabidopsis stigmas intersect at Exo70A1, a putative component of the exocyst complex. Plant Cell 21: 2655–2671 PubMed PMC

Samuels A. (1993) The effects of silicon supplementation on cucumber fruit: changes in surface characteristics. Ann Bot 72: 433–440

Samuels AL, Glass ADM, Ehret DL, Menzies JG (1991a) Mobility and deposition of silicon in cucumber plants. Plant Cell Environ 14: 485–492

Samuels AL, Glass ADM, Ehret DL, Menzies JG (1991b) Distribution of silicon in cucumber leaves during infection by powdery mildew fungus (Sphaerotheca fuliginea). Can J Bot 69: 140–146

Sangster AG, Hodson MJ (2007) Silica in higher plants. In D Evered, M O'Connor, eds, Novartis Foundation Symposia: Silicon Biochemistry. John Wiley & Sons, Chichester, UK, pp 90–111

Schindelin J, Arganda-Carreras I, Frise E, Kaynig V, Longair M, Pietzsch T, Preibisch S, Rueden C, Saalfeld S, Schmid B, et al. (2012) Fiji: an open-source platform for biological-image analysis. Nat Methods 9: 676–682 PubMed PMC

Sekereš J, Pejchar P, Šantrůček J, Vukašinović N, Žárský V, Potocký M (2017) Analysis of exocyst subunit EXO70 family reveals distinct membrane polar domains in tobacco pollen tubes. Plant Physiol 173: 1659–1675 PubMed PMC

Seo DH, Ahn MY, Park KY, Kim EY, Kim WT (2016) The N-terminal UND motif of the Arabidopsis U-box E3 ligase PUB18 is critical for the negative regulation of ABA-mediated stomatal movement and determines its ubiquitination specificity for exocyst subunit Exo70B1. Plant Cell 28: 2952–2973 PubMed PMC

Sinlapadech T, Stout J, Ruegger MO, Deak M, Chapple C (2007) The hyper-fluorescent trichome phenotype of the brt1 mutant of Arabidopsis is the result of a defect in a sinapic acid: UDPG glucosyltransferase. Plant J 49: 655–668 PubMed

Stegmann M, Anderson RG, Ichimura K, Pecenkova T, Reuter P, Žársky V, McDowell JM, Shirasu K, Trujillo M (2012) The ubiquitin ligase PUB22 targets a subunit of the exocyst complex required for PAMP-triggered responses in Arabidopsis. Plant Cell 24: 4703–4716 PubMed PMC

Stegmann M, Anderson RG, Westphal L, Rosahl S, McDowell JM, Trujillo M (2013) The exocyst subunit Exo70B1 is involved in the immune response of Arabidopsis thaliana to different pathogens and cell death. Plant Signal Behav 8: e27421. PubMed PMC

Sunkar R, Zhu J-K (2004) Novel and stress-regulated microRNAs and other small RNAs from Arabidopsis. Plant Cell 16: 2001–2019 PubMed PMC

Tian J, Han L, Feng Z, Wang G, Liu W, Ma Y, Yu Y, Kong Z (2015) Orchestration of microtubules and the actin cytoskeleton in trichome cell shape determination by a plant-unique kinesin. eLife 4: e09351 PubMed PMC

Tihlaříková E, Neděla V, Shiojiri M (2013) In situ study of live specimens in an environmental scanning electron microscope. Microsc Microanal 19: 914–918 PubMed

Töller A, Brownfield L, Neu C, Twell D, Schulze-Lefert P (2008) Dual function of Arabidopsis glucan synthase-like genes GSL8 and GSL10 in male gametophyte development and plant growth. Plant J 54: 911–923 PubMed

Verma DP, Hong Z (2001) Plant callose synthase complexes. Plant Mol Biol 47: 693–701 PubMed

Vivancos J, Labbé C, Menzies JG, Bélanger RR (2015) Silicon-mediated resistance of Arabidopsis against powdery mildew involves mechanisms other than the salicylic acid (SA)-dependent defence pathway. Mol Plant Pathol 16: 572–582 PubMed PMC

Vogel J, Somerville S (2000) Isolation and characterization of powdery mildew-resistant Arabidopsis mutants. Proc Natl Acad Sci USA 97: 1897–1902 PubMed PMC

Vukašinović N, Oda Y, Pejchar P, Synek L, Pečenková T, Rawat A, Sekereš J, Potocký M, Žárský V (2017) Microtubule-dependent targeting of the exocyst complex is necessary for xylem development in Arabidopsis. New Phytol 213: 1052–1067 PubMed

Waterkeyn L, Dupont C (1982) L’observation des dépôts pariétaux de silice au microscope électronique à balayage. Bulletin Societe Royale de Botanique de Belgique 115: 156–160

Webster TR. (1992) Developmental problems in Selaginella (Selaginellaceae) in an evolutionary context. Ann Mo Bot Gard 79: 632–647

Wu H, Turner C, Gardner J, Temple B, Brennwald P (2010) The Exo70 subunit of the exocyst is an effector for both Cdc42 and Rho3 function in polarized exocytosis. Mol Biol Cell 21: 430–442 PubMed PMC

Yi X, Zhang Z, Ling Y, Xu W, Su Z (2015) PNRD: a plant non-coding RNA database. Nucleic Acids Res 43: D982–D989 PubMed PMC

Yue P, Zhang Y, Mei K, Wang S, Lesigang J, Zhu Y, Dong G, Guo W (2017) Sec3 promotes the initial binary t-SNARE complex assembly and membrane fusion. Nat Commun 8: 14236. PubMed PMC

Zárský V, Cvrcková F, Potocký M, Hála M (2009) Exocytosis and cell polarity in plants - exocyst and recycling domains. New Phytol 183: 255–272 PubMed

Zhang C, Wang L, Zhang W, Zhang F (2013) Do lignification and silicification of the cell wall precede silicon deposition in the silica cell of the rice (Oryza sativa L.) leaf epidermis? Plant Soil 372: 137–149

Zhao T, Rui L, Li J, Nishimura MT, Vogel JP, Liu N, Liu S, Zhao Y, Dangl JL, Tang D (2015) A truncated NLR protein, TIR-NBS2, is required for activated defense responses in the exo70B1 mutant. PLoS Genet 11: e1004945. PubMed PMC

Zhao Y, Liu J, Yang C, Capraro BR, Baumgart T, Bradley RP, Ramakrishnan N, Xu X, Radhakrishnan R, Svitkina T, Guo W (2013) Exo70 generates membrane curvature for morphogenesis and cell migration. Dev Cell 26: 266–278 PubMed PMC

Interplay of EXO70 and MLO proteins modulates trichome cell wall composition and susceptibility to powdery mildew

Plasma membrane phospholipid signature recruits the plant exocyst complex via the EXO70A1 subunit

Functional Specialization within the EXO70 Gene Family in Arabidopsis

Synergy among Exocyst and SNARE Interactions Identifies a Functional Hierarchy in Secretion during Vegetative Growth

Redundant and Diversified Roles Among Selected Arabidopsis thaliana EXO70 Paralogs During Biotic Stress Responses

Regulation of Exocyst Function in Pollen Tube Growth by Phosphorylation of Exocyst Subunit EXO70C2

Arabidopsis Trichome Contains Two Plasma Membrane Domains with Different Lipid Compositions Which Attract Distinct EXO70 Subunits

In-situ preparation of plant samples in ESEM for energy dispersive x-ray microanalysis and repetitive observation in SEM and ESEM